Skip to main content
SpringerLink
Log in

Enhancing the Prebiotic Relevance of a Set of Covalently Self-Assembling, Autorecombining RNAs Through In Vitro Selection

  • Published:
Journal of Molecular Evolution Aims and scope Submit manuscript

Abstract

An in vitro form of the self-splicing group-I intron interrupting the Azoarcus tRNAIle was shortened by ~10% with the removal of helix P6a. This deletion reduced the reverse-splicing activity of the ribozyme about 10-fold. Through in vitro selection, this activity was restored in several low-error mutants. A number of mutations were found that improved reverse-splicing activity through both increased k obs and better folding. The deletion mutant could be fragmented into as many as three discrete pieces, which, when incubated together, were capable of covalent self-assembly through energy-neutral transesterification reactions, a process called autorecombination. A subset of the mutations identified through in vitro selection for reverse-splicing were exaptations in that they were also shown to augment the autorecombination reactions, leading to higher yields of covalently self-assembled products, making this the smallest such system yet discovered.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  • Basu S, Rambo RP, Strauss-Soukup J, Cate JH, Ferré-D’Amaré AR, Strobel SA, Doudna JA (1998) A specific monovalent metal ion integral to the AA platform of the RNA tetraloop receptor. Nat Struct Biol 5:986–992

    Article  CAS  PubMed  Google Scholar 

  • Bell MA, Johnson AK, Testa SM (2002) Ribozyme-catalyzed excision of targeted sequences from within RNAs. Biochemistry 41:15327–15333

    Article  CAS  PubMed  Google Scholar 

  • Boussau B, Blanquart S, Necsulea A, Lartillot N, Gouy M (2008) Parallel adaptations to high temperatures in the Archaean eon. Nature 456:942–945

    Article  CAS  PubMed  Google Scholar 

  • Burke DH, Willis JH (1998) Recombination, RNA evolution, and bifunctional RNA molecules isolated through chimeric SELEX. RNA 4:1165–1175

    Article  CAS  PubMed  Google Scholar 

  • Burton AS, Lehman N (2006) Calcium(II)-dependent catalytic activity of the Azoarcus ribozyme: testing the limits of resolution for in vitro selection. Biochimie 88:819–825

    Article  CAS  PubMed  Google Scholar 

  • Burton AS, Madix RA, Vaidya N, Riley CA, Hayden EJ, Chepetan A, Dìaz Arenas C, Larson BC, Lehman N (2009) Gel purification of radiolabeled nucleic acids via phosphorimaging: Dip-N-Dot. Anal Biochem 388:351–352

    Article  CAS  PubMed  Google Scholar 

  • Cadwell RC, Joyce GF (1992) Randomization of genes by PCR mutagenesis. PCR Methods Appl 2:28–33

    CAS  PubMed  Google Scholar 

  • Dotson PP, Johnson AK, Testa SM (2008) Tetrahymena thermophila and Candida albicans group I intron-derived ribozymes can catalyze the trans-excision-splicing reaction. Nucleic Acids Res 36:5281–5289

    Article  CAS  PubMed  Google Scholar 

  • Eigen M (1971) Selforganization of matter and the evolution of biological macromolecules. Naturwissenschaften 58:465–523

    Article  CAS  PubMed  Google Scholar 

  • Engels J, Uhlmann E (1988) Gene synthesis. Adv Biochem Eng Biotechnol 37:73–127

    CAS  PubMed  Google Scholar 

  • Ferris JP (2002) Montmorillonite catalysis of 30–50 mer oligonucleotides: laboratory demonstration of potential steps in the origin of the RNA world. Orig Life Evol Biosph 32:311–332

    Article  CAS  PubMed  Google Scholar 

  • Gesteland R, Cech TR, Atkins J (eds) (2006) The RNA World, 3rd edn. Cold Spring Harbor Press, New York

    Google Scholar 

  • Gilbert W (1986) The RNA world. Nature 319:618

    Article  Google Scholar 

  • Gould SJ, Vrba ES (1982) Exaptation-A missing term in the science of form. Paleobiology 8:4–15

    Google Scholar 

  • Hayden EJ, Lehman N (2006) Self-assembly of a group I intron from inactive oligonucleotide fragments. Chem Biol 13:909–918

    Article  CAS  PubMed  Google Scholar 

  • Hayden EJ, Riley CA, Burton AS, Lehman N (2005) RNA-directed construction of structurally complex and active ligase ribozymes through recombination. RNA 11:1678–1687

    Article  CAS  PubMed  Google Scholar 

  • Huang Z, Szostak J (1996) A simple method for 3′-labeling of RNA. Nucl Acids Res 24:4360–4361

    Article  CAS  PubMed  Google Scholar 

  • Huang Z, Pei W, Han Y, Jayaseelan S, Shekhtman A, Shi H, Niu L (2009) One RNA aptamer sequence, two structures: a collaborating pair that inhibits AMPA receptors. Nucl Acids Res 37:4022–4032

    Article  CAS  PubMed  Google Scholar 

  • Johnston WK, Unrau PJ, Lawrence MS, Glasner ME, Bartel DP (2001) RNA-catalyzed RNA polymerization: accurate and general RNA-templated primer extension. Science 292:1319–1325

    Article  CAS  PubMed  Google Scholar 

  • Joyce GF (1991) The rise and fall of the RNA world. New Biol 3:399–407

    CAS  PubMed  Google Scholar 

  • Joyce GF (1998) Nucleic acid enzymes: playing with a fuller deck. Proc Natl Acad Sci USA 95:5845–5847

    Article  CAS  PubMed  Google Scholar 

  • Joyce GF (2002) The antiquity of RNA-based evolution. Nature 418:214–221

    Article  CAS  PubMed  Google Scholar 

  • Kuhn H (1972) Self-organization of molecular systems and evolution of the genetic apparatus. Angew Chem Int Ed Engl 11:798–820

    Article  CAS  PubMed  Google Scholar 

  • Lehman N (2003) A case for the extreme antiquity of recombination. J Mol Evol 56:770–777

    Article  CAS  PubMed  Google Scholar 

  • Lehman N (2008) A recombination-based model for the origin and early evolution of genetic information. Chem Biodivers 5:1707–1717

    Article  CAS  PubMed  Google Scholar 

  • Lehman N, Joyce GF (1993) Evolution in vitro of an RNA enzyme with altered metal dependence. Nature 361:182–185

    Article  CAS  PubMed  Google Scholar 

  • McGinness KE, Joyce GF (2003) In search of an RNA replicase ribozyme. Chem Biol 10:5–14

    Article  CAS  PubMed  Google Scholar 

  • Orgel LE (1986) RNA catalysis and the origins of life. J Theor Biol 123:127–149

    Article  CAS  PubMed  Google Scholar 

  • Pan J, Woodson SA (1998) Folding intermediates of a self-splicing RNA: mispairing of the catalytic core. J Mol Biol 280:597–609

    Article  CAS  PubMed  Google Scholar 

  • Pan J, Deras ML, Woodson SA (2000) Fast folding of a ribozyme by stabilizing core interactions: evidence for multiple folding pathways in RNA. J Mol Biol 296:133–144

    Article  CAS  PubMed  Google Scholar 

  • Rich A (1962) On the problems of evolution and biochemical information transfer. In: Kasha M, Pullmann B (eds) Horizons in biochemistry. Academic Press, New York, pp 103–126

    Google Scholar 

  • Riley CA, Lehman N (2003) Generalized RNA-directed recombination of RNA. Chem Biol 10:1233–1243

    Article  CAS  PubMed  Google Scholar 

  • Rydzanicz R, Zhao XS, Johnson PE (2005) Assembly PCR oligo maker: a tool for designing oligodeoxynucleotides for constructing long DNA molecules for RNA production. Nucleic Acids Res 33:W521–W525

    Article  CAS  PubMed  Google Scholar 

  • Schmitt T, Lehman N (1999) Non-unity molecular heritability demonstrated by continuous evolution in vitro. Chem Biol 6:857–869

    Article  CAS  PubMed  Google Scholar 

  • Sclavi B, Sullivan M, Chance MR, Brenowitz M, Woodson SA (1998) RNA folding at millisecond intervals by synchrotron hydroxyl radical footprinting. Science 279:1940–1943

    Article  CAS  PubMed  Google Scholar 

  • Striggles JC, Martin MB, Schmidt FJ (2006) Frequency of RNA–RNA interaction in a model of the RNA World. RNA 12:353–359

    Article  CAS  PubMed  Google Scholar 

  • Szostak J, Ellington A (1993) In vitro selection of functional RNA sequences. In: Gesteland R, Atkins J (eds) The RNA world, 1st edn. Cold Spring Harbor Press, New York, pp 511–533

    Google Scholar 

  • Treiber DK, Williamson JR (2001) Beyond kinetic traps in RNA folding. Curr Opin Struct Biol 11:309–314

    Article  CAS  PubMed  Google Scholar 

  • Vartanian JP, Henry M, Wain-Hobson S (1996) Hypermutagenic PCR involving all four transitions and a sizeable proportion of transversions. Nucleic Acids Res 24:2627–2631

    Article  CAS  PubMed  Google Scholar 

  • Zaher HS, Unrau PJ (2007) Selection of an improved RNA polymerase ribozyme with superior extension and fidelity. RNA 13:1017–1026

    Article  CAS  PubMed  Google Scholar 

  • Zarrinkar P, Williamson JR (1994) Kinetic intermediates in RNA folding. Science 265:918–924

    Article  CAS  PubMed  Google Scholar 

  • Zhang L, Xiao M, Lu C, Zhang Y (2005) Fast formation of the P3–P7 pseudoknot: a strategy for efficient folding of the catalytically active ribozyme. RNA 11:59–69

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The authors thank N. Vaidya for technical assistance and E. Hayden for useful discussions. This study was supported by NASA Exobiology Grant number NNX07AU05G to N. Lehman.

Author information

Authors and Affiliations

  1. Department of Chemistry, Portland State University, P.O. Box 751, Portland, OR, 97207, USA

    Aaron S. Burton & Niles Lehman

Authors
  1. Aaron S. Burton
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Niles Lehman
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Niles Lehman.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Burton, A.S., Lehman, N. Enhancing the Prebiotic Relevance of a Set of Covalently Self-Assembling, Autorecombining RNAs Through In Vitro Selection. J Mol Evol 70, 233–241 (2010). https://doi.org/10.1007/s00239-010-9325-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00239-010-9325-3

Keywords

Search

Navigation